This project tests an improved DC vaccine which is designed to promote in vivo cross presentation and determinant spreading. Based on results from our previous trials, we have made several important improvements: engineering the DC with 3 full-length, defined, tumor antigens to activate multiple CD8* and CD4'^ T cell clones (reducing the concern for antigen loss variants); providing antigen presentation for the life of the DC; providing cognate CD4* T cell help to the CD8+ T cells (helped CTL); using a matured DC (a cocktail specifically matched to adenovirus (AdV) transduction signals); activating innate immunity via NK cell migration and activation; and boosting with systemic IFN alpha (for endogenous DC type 1 skewing, improved cross-priming and direct effects on T cells). Together, this vaccine strategy should more potently activate a polyclonal anti-tumor response incorporating multiple adaptive and innate effectors which we predict will lead to a higher frequency of patients who not only activate vaccine-encoded antigen-specific T cell responses, but also develop determinant spreading and a significant clinical response. The three aims are: Aim 1: Completion of the AdVTMM2/DC +/- IFN alpha clinical trial 1.a. Clinical outcomes in the stage IV patients; 1.b. Immunologic outcomes (from blood and TIL; baseline, post-vaccine and post-IFN alpha);1.c. AdV-specific cellular and humoral responses Aim 2: Mechanism and biomarkers of clinical response 2.a. DC Vaccine Transcriptional Profiling (+/- maturation, +/- AdVTMM2); 2.b. Tumor Transcriptional Profiling (baseline, post-vaccine and post-IFN alpha); 2.c. Peripheral blood signaling induced by IFN alpha (STATs); 2.d. Serum profiling (autoimmunity antibodies, LDH, CRP cytokines); 2.e. Molecular mimicry with mycoplasma (impact of baseline memory to mycoplasma); 2.f. Germline DNA SNP analysis Aim 3: Mechanism of NK cell activation in anti-melanoma immunity In vitro studies to expand our recent findings in AdV/DC-NK cell cross-talk with banked melanoma patient blood and tumor (primary tumor expanded to cell lines): 3.a. Direct NK cell interactions with melanoma tumor cells (cytotoxicity, cytokines); 3b: Helper/type 1 skewing role in shaping adaptive CDS* and CD4* T cell responses; 3c: Melanoma tumor impact on NK cell function.